Liver is a glandular tissue connected to digestive tract and has various important functions for living organisms, such as production and secretion of bile, a detoxification action, carbohydrate metabolism, protein metabolism, production of a blood coagulation factor, a hormone regulation action, and storage of various components of living organisms, including fat, glycogen, a protein, and a vitamin. Therefore, when those functions are acutely or chronically damaged due to viral infection, a drug or a toxic agent, or excess alcohol intake, the maintenance of homeostasis of liver functions is disrupted, resulting in a critical health problem. Among various diseases found in comprehensive medical examination or the like, liver dysfunction accounts for a high proportion of the diseases, and about 30% of Japanese adults are estimated to have liver dysfunction.
Diseases included in liver disorders are classified based on their etiologies or clinical symptoms. For example, the diseases may be classified based on their etiologies into viral hepatitis, drug-induced liver injury, alcoholic liver injury, autoimmune liver injury, metabolic liver injury, and the like. In addition, the diseases may be classified based on their clinical symptoms into hepatocellular injury type and cholestasis type. Hepatocellular injury is found in viral hepatitis, toxic liver injury, fatty liver, cirrhosis, and the like. Hepatocellular injury is caused by, for example, necrosis, fatty metamorphosis, multinucleation, or nuclear vacuolar degeneration of a hepatocyte. The hepatocyte, which is also referred to as hepatic parenchymal cell, is one of the cells constituting liver and accounts for most major parts of liver. The hepatocyte is an exocrine cell which secrets bile, while it is also an endocrine cell which secretes plasma proteins into an apical membrane side and stores glycogen to regulate blood glucose. Therefore, when the hepatocyte is damaged, a significant reduction in liver function is caused. Cholestasis is caused by blockade of bile excretion from liver due to some cause, with the result that abnormal excretion of bile is brought about over a part or whole of intrahepatic or extrahepatic bile duct system, and thereby the bile accumulates in liver and blood. Then, various symptoms such as jaundice and hepatitis are induced.
At present, diagnosis and treatment of liver diseases are performed according to an approach in which a physician empirically diagnoses a liver injury type using a symptom of a patient and values of serum biochemical parameters as indicators and then treatment is started (Non Patent Literature 1). Enzymes which leak from the hepatocyte by hepatocellular injury are used as the biochemical parameters. Specifically, there are used, for example, aspartate aminotransferase (hereinafter abbreviated as AST; sometimes referred to as glutamate-oxaloacetate transaminase (GOT)), alanine aminotransferase (hereinafter abbreviated as ALT; sometimes referred to as glutamate-pyruvate transaminase (GPT)), γ-glutamyl transpeptidase (hereinafter abbreviated as γ-GTP), and alkaline phosphatase (hereinafter abbreviated as ALP).
In addition, in non-clinical and clinical practices, measurement of total bile acids (hereinafter abbreviated as TBA) is generally performed. Bile acids is a general term for compounds that are steroid derivatives having cholanic acid skeletons widely found in bile of mammals and are major components of bile which plays a key role in digestion and absorption of fat. The bile acids are produced from metabolism of cholesterol in hepatocytes of liver (FIG. 1), followed by conjugating with glycine and taurine (aminoethylsulfonic acid) or the like, and then excreted as bile. The bile acids are roughly classified into primary bile acids and secondary bile acids. The primary bile acids are bile acids which are synthesized in liver. Examples of the primary bile acids may include cholic acid (hereinafter sometimes abbreviated as CA), chenodeoxycholic acid (hereinafter sometimes abbreviated as CDCA), and ursodeoxycholic acid (hereinafter sometimes abbreviated as UDCA) which has an isomeric relationship with CDCA. The secondary bile acids are bile acids which are produced from the primary bile acids excreted via bile duct to intestinal tract in a dehydroxylation reaction, a dehydrogenation reaction, a hydrogenation reaction, or a deconjugation reaction by enterobacteria. Examples of the secondary bile acids may include deoxycholic acid (hereinafter sometimes abbreviated as DCA) and LCA. Those five kinds of bile acid components given as examples each have three forms which are a free form, a glycine-conjugated form, and a taurine-conjugated form. In addition, some bile acid components each have a sulfated form or a glucuronidated form. In many animal species including humans, CA and CDCA having hydroxyl groups at position 3, 7, and 12 of a cholanic acid skeleton are produced as the primary bile acids. Meanwhile, some animals produce bile acids inherent in the species. For example, α-muricholic acid (hereinafter sometimes abbreviated as αMCA) and β-muricholic acid (hereinafter sometimes abbreviated as βMCA) are found in mice and hyocholic acid (hereinafter sometimes abbreviated as HCA) is found in swine, and these bile acids are rare in humans and are characteristic of each of the animal species. αMCA and βMCA are primary bile acids and taurine-conjugated forms thereof are also known. HCA is a primary bile acid and produces hyodeoxycholic acid (hereinafter abbreviated as HDCA) by its 7α-dehydroxylation in intestine. Those bile acid components also have glycine-conjugated forms and taurine-conjugated forms.
In treatment of liver diseases, discrimination of a liver injury type is important for selection of treatment suitable for each liver injury type. Hitherto, liver injury types have been determined based on biochemical parameters. For example, a liver injury type is determined using, as indicators, increases in ALT and AST in hepatocellular injury type and increases in ALP and γ-GTP in cholestasis type. However, a determination method based on biochemical parameters sometimes leads to misjudgment of a liver injury type. Therefore, an attempt has been made to measure levels of many components constituting bile acids in serum and discriminate a liver injury type based on the results to further determine a therapeutic strategy.
There have been previous reports on various relationships between classification of liver injury types based on etiologies and levels of bile acid components (Non Patent Literatures 2 to 6). However, no consensus has been reached and no clinical application has been achieved.
In addition, there have been some reports on the association between classification of liver injury types based on clinical symptoms and levels of bile acid components (Non Patent Literatures 6 to 8). However, similarly to the classification of liver injury types based on etiologies, a variety of results have been shown and no consensus has been reached. Meanwhile, for classification of types of drug-induced liver injury based on clinical symptoms, there has been proposed an assessment method using biochemical parameters rather than bile acid components (Non Patent Literature 9). Based on this proposal, the Japan Society of Hepatology either proposes scoring of drug-induced liver injury (Non Patent Literature 10). Further, the inventors of the present invention have reported that a liver disease caused by administration of a drug to a rat was compared to a typical cholestasis model obtained by ligation of bile duct, or administration of 1-naphtyl isothiocyanate (ANIT), or the like, and that injury types of liver diseases may be discriminated by measuring levels of bile acid components (Non Patent Literature 11). However, at present, there is no report on a definitive biomarker that clinically diagnoses an early stage of cholestasis in liver at a capillary bile duct level. Thus, the decision of a therapeutic strategy is difficult in many cases.